A family of viruses, named Circoviridae, found in a range of plant and animal species and commonly referred to as circoviruses, are characterized as round, non-enveloped virions with mean diameters from 17 to 23.5 nm containing circular, single-stranded deoxyribonucleic acid (ssDNA). The ssDNA genome of the circoviruses represent the smallest viral DNA replicons known. As disclosed in WO 99/45956, at least six viruses have been identified as members of the family according to The Sixth Report of the International Committee for the Taxonomy of Viruses (Lukert, P. D. et al. 1995, The Circoviridae, pp. 166–168. In F. A. Murphy, et al. (eds.) Virus Taxonomy, Sixth Report of the International Committee on Taxonomy of Viruses, Arch. Virol. 10 Suppl.).
Animal viruses included in the family are chicken anemia virus (CAV); beak and feather disease virus (BFDV); porcine circovirus (PCV); and pigeon circovirus. PCV was originally isolated in porcine kidney cell cultures. PCV replicates in the cell nucleus and produces large intranuclear inclusion bodies. See Murphy et al. (1999, Circoviridae p. 357–361, Veterinary Virology, 3rd ed. Academic Press, San Diego). There are currently two recognized types of PCV, PCV type 1 (PCV1) and PCV type 2 (PCV2). PCV1, isolated as a persistent contaminant of the continuous porcine kidney cell line PK-15 (ATCC CCL31), does not cause detectable cytopathic effects in cell culture and fails to produce clinical disease in pigs after experimental infection (see Allan G., 1995, Vet. Microbiol. 44: 49–64; Tischer, I. et al., 1982, Nature 295:64–66; and Tischer, I. et al., 1986, Arch. Virol. 91:271–276). PCV2, in contrast to PCV1, is closely associated with post weaning multisystemic wasting syndrome (PMWS) in weanling pigs (see Allan G. et al., 1998, Europe. J. Vet. Diagn. Investig. 10:3–10; Ellis, J. et al., 1998, Can. Vet. J. 39:44–51 and Morozov, I. et al., 1998, J. Clin. Microbiol. 36:2535–2541). The nucleotide sequences for PCV1 are disclosed in Mankertz, A., et al. (1997, J. Virol. 71:2562–2566) and Meehan, B. M., et al. (1997, J. Gen. Virol. 78:221–227) and the nucleotide sequences for PCV2 are disclosed in Hamel, A. L. et al. (1998, J. Virol. 72:5262–5267); Mankertz, A. et al. (2000, Virus Res. 66:65–77) and Meehan, B. M. et al. (1998, J. Gen. Virol. 79:2171–2179). Strains of PCV2 are disclosed in WO 00/01409 and have been deposited at the European Collection of Cell Cultures, Centre for Applied Microbiology & Research, Porton Down, Salisbury, Wiltshire SP4 OJG, United Kingdom and include: accession No. V97100219; accession No. V9700218; accession No. V97100217; accession No. V98011608; and accession No. V98011609. WO 00/77216 also discloses PCV2.
Published studies to date on PCV2 used either tissue homogenate or cultured virus derived from field isolates. Tischer et al. (1987, Arch Virol. 96:39–57) report that porcine kidney cells are stimulated to entry to the S phase in the cell cycle by D-glucosamine treatment. However, the treatment must be performed with caution because D-glucosamine is toxic for cell culture (see, Allan et al., (2000). J. Vet. Diagn. Investigation. 12:3–14). There remains a need for methods for culturing circovirus, such as for example, PCV1 and PCV2, and other circoviruses, such that pure circovirus is obtained. Such methods would be advantageous, in particular for preparation of PCV2 antigens as vaccines directed against PMWS. The present invention addresses that need.
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